Silybin A and B were separated from commercial silibinin using the preparative HPLC method. The described method is rapid and effective in obtaining gram-scale amounts of two diastereo-isomers with minimal effort. In our approach, silibinin was dis-solved in THF (solubility greater than 100 mg/mL), an alternative solvent to H 2 O or MeOH in which silibinin has a very low solubil-ity (ca 0.05–1.5 mg/mL), and then separated into its two compo-nents using preparative RP-HPLC. By starting with purified dia-stereoisomers, it was possible to obtain the two enantiomers of 2,3-dehydrosilybin in good yields and optically pure using an ef-ficient oxidation procedure. All of the purified products were fully characterised using NMR (1 H, 13 C), CD, [α] D , and ESI MS anal-yses. The purities of the products, which were evaluated using analytical HPLC, were greater than 98 % in all cases. Key words prep‑RP‑HPLC · flavonolignans · Silybum marianum · Astera-ceae · silibinin · silybin A · silybin B · 2,3‑dehydrosilybin